Aromatase-inhibiting composition containing azole derivative

ABSTRACT

The disclosure describes a pharmaceutical composition comprising an aromatase-inhibiting effective amount of a compound selected from the group consisting of compounds of the formula (I): ##STR1## including their stereoisomers and salts thereof, wherein R 1  is halogen or phenyl; m is 0, 1, 2 or 3; k and n each are independently 0, 1 or 2; R 2  and R 3  each are independently H or OH; R 6  and R 7  each are independently H or C 1-4  alkyl; R 8  and R 9  each are H or R 8  and R 9  form --C(R 4 ) (R 5 )--C(R 11 ) (R12)-- bond wherein R 4 , R 5 , R 11  and R 12  each are independently H or C 1-4  alkyl; Y is N or CH; and R 10  is H or halogen, or R 10  combines with R 2  to form --O-- bond, with proviso that if R 8  and R 9  form --C (R 4 ) (R 5 )--C(R 11 ) (R 12 )-- bond R 3 , R 6  and R 7  each are H, and k and n each are 1, then R 2  and R 10  form --O-- bond, and a pharmaceutically acceptable carrier or diluent.

This is a Division of application Ser. No. 08/325,161 filed Oct. 21,1994.

BACKGROUND OF THE INVENTION

The present invention relates to an aromatase-inhibiting compositioncontaining an azole derivative.

Estrogen, a sex hormone, is synthesized from androgenic steroids in thetissues such as ovary or placenta. In the biosynthetic pathway for theestrogen formation from an androgenic steroid, aromatization isessential, and aromatase takes part in such aromatization. It istherefore expected that if the aromatase can be effectively inhibited,estrogen-dependent diseases can be effectively treated (Cancer Research,Vol. 42, Suppl. 8, 3261s, 1982), and to this end certain aromataseinhibitors have been proposed.

4-Hydroxyandrostenedione is known as such an aromatase inhibitor.However, many of the steroid type drugs are generally attended withconsiderable side effects in practical use.

It is demanded to provide an aromatase inhibitor possessing an excellentinhibitory activity on the aromatase.

As a result of the present inventors' earnest studies, it has been foundthat azole derivatives having a non-steroidal chemical structure quitedifferent from 4-hydroxyandrostenedione have an inhibitory activity onthe aromatase. The present invention has been attained on the basis ofthis finding.

SUMMARY OF THE INVENTION

An object of the present invention is to provide an aromatase inhibitingcomposition containing an azole derivative which is low in toxicity(side effects) and shows a remarkable inhibitory activity on thearomatase.

To achieve the aim, in an aspect of the present invention, there isprovided a pharmaceutical composition comprising an aromatase-inhibitingeffective amount of a compound selected from the group consisting ofcompounds of the formula (I): ##STR2## including their stereoisomers andsalts thereof, wherein R¹ is halogen or phenyl; m is 0, 1, 2 or 3; k andn each are independently 0, 1 or 2; R² and R³ each are independently Hor OH; R⁶ and R⁷ each are independently H or C₁₋₄ alkyl; R⁸ and R⁹ eachare H or R⁸ and R⁹ form --C(R⁴)(R⁵)--C(R¹¹)(R¹²)-- bond wherein R⁴, R⁵,R¹¹ and R¹² each are independently H or C₁₋₄ alkyl; Y is N or CH; andR¹⁰ is H or halogen, or R¹⁰ combines with R² to form --O-- bond, withproviso that if R⁸ and R⁹ form --C(R⁴)(R⁵)--C(R¹¹)(R¹²)-- bond, R³, R⁶and R⁷ each are H, and k and n each are 1, then R² and R¹⁰ form --O--bond, and a pharmaceutically acceptable carrier or diluent.

DETAILED DESCRIPTION OF THE INVENTION

The azole derivative according to the present invention is preferably anisolated single stereoisomer. The "stereoisomers" referred to hereininclude geometrical isomers, optical isomers and diastereoisomers.

The preferred combinations of the substituents in the above formula areas follows: R¹ is Cl, F or phenyl; m is 0, 1 or 2; k and n each areindependently 0 or 1; R² and R³ are as defined above; R⁶ and R⁷ each areindependently H or C₁₋₂ alkyl; R⁸ and R⁹ form --C(R⁴) (R⁵)--C(R¹¹)(R¹²)-- bond, wherein R⁴, R⁵, R¹¹ and R¹² each are independently H orC₁₋₂ alkyl; and Y and R¹⁰ are as defined above.

The more preferred combinations of the substituents are as specifiedbelow in (1) to (4).

(1) R¹ is Cl or Br; m is 0, 1 or 2; k and n each are independently 0 or1; R², R⁸, R⁹ and R¹⁰ each are H; R³ is OH; R⁶ and R⁷ each areindependently C₁₋₂ alkyl; and Y is N or CH.

(2) R¹ is Cl, Br or phenyl; m is 0, 1 or 2; k and n each areindependently 0 or 1; R² and R¹⁰ each are H; R³ is OH; R⁶ and R⁷ eachare independently H or C₁₋₂ alkyl; R⁸ and R⁹ form --CH₂ --CH₂ --bond;and Y is N or CH.

(3) R¹ is Cl or Br; m is 0, 1 or 2; k and n each are independently 0 or1; R³, R⁶ and R⁷ each are H; R⁸ and R⁹ form --CH₂ --C(R¹¹) (R¹²)-- bond,wherein R¹¹ and R¹² each are independently H or C₁₋₂ alkyl; R¹⁰ and R²form --O-- bond; and Y is N or CH.

(4) R¹ is Cl or Br; m is 0, 1 or 2; k is 1; n is 0 or 1; R³, R⁶ and R⁷each are H; R² is OH; R⁸ and R⁹ form --C(R⁴)(R⁵)--CH₂ -- bond, whereinR⁴ and R⁵ each are independently C₁₋₂ alkyl; Y is N or CH; and R¹⁰ is Hor halogen, or R¹⁰ combines with R² to form --O-- bond.

The azole derivatives according to the present invention can be producedfrom the following processes.

Process A

In case no ring is formed by R⁸ and R⁹, and R² and R¹⁰ :

A preparation process of a compound of formula (I) is shown in Scheme 1.##STR3##

Step (a)

A compound of formula (IX) wherein R², R⁶, R⁷, R⁸ and R⁹ are as definedabove; R¹³ is C₁₋₃ alkyl; and n is 1 or 2, is reacted with ethyleneglycol at 60° to 140° C. in the presence of p-toluenesulfonic acid toform a compound of formula (VIII) wherein R², R⁶, R⁷, R⁸, R⁹, n and R¹³are as defined above.

Step (b)

The compound of formula (VIII) is reacted with LiAlH₄ at -10° to +35° C.to form a compound of formula (VII) wherein R², R⁶, R⁷, R⁸, R⁹ and n areas defined above.

Step (c)

The compound of formula (VII) is reacted with methanesulfonyl chlorideat -10° to +10° C. to form a compound of formula (VI) wherein R², R⁶,R⁷, R⁸, R⁹ and n are as defined above.

Step (d)

The compound of formula (VI) is reacted with a compound of formula (V)wherein M is H or a metal; and Y is N or CH, at 40° to 100° C. to form acompound of formula (IV) wherein R², R⁶, R⁷, R⁸, R⁹, n and Y are asdefined above.

Step (e)

The compound of formula (IV) is deketalated with an acid at 10° to 80°C. to form a compound of formula (III) wherein R², R⁶, R⁷, R⁸, R⁹, Y andn are as defined above.

Step (f)

The compound of formula (III) is reacted with a compound of formula (II)wherein X is halogen; R¹⁰ is H or halogen; R¹ is halogen or phenyl; m is0, 1, 2 or 3; and k is 0, 1 or 2, at -80° to +80° C. to obtain acompound of formula (I).

Process B-1

In case R⁸ and R⁹ form --C(R⁴) (R⁵)--C(R¹¹) (R¹²)-- bond and a phenylderivative is used in the reaction:

The preparation steps are shown in Scheme 2. ##STR4##

A compound of formula (II) wherein X is halogen, and k, m, R¹ and R¹⁰are as defined above, is added to an ether solution of magnesium. To thesolution, a solution of a compound of formula (III) wherein R², R⁴, R⁵,R⁶, R⁷, R¹¹, R¹², n and Y are as defined above, is added dropwise andreacted at 5° to 20° C. to form a compound of formula (I). The compoundof formula (III) can be derived from cyclopentanone.

Process B-2

In case R⁸ and R⁹ form --C(R⁴)(R⁵)--C(R¹¹)(R¹²)-- bond and an azolederivative is used in the reaction:

The preparation steps are shown in Scheme 3. ##STR5##

A compound of formula (III') wherein R¹, R³, R⁴, R⁵, R⁶, R⁷, R¹⁰, R¹¹,R¹², m and k are as defined above, is reacted with a compound of formula(V) wherein M and Y are as defined above, to form a compound of formula(I). The compound of formula (I) is reacted with a base, or the reactionis further advanced to obtain a compound of formula (I').

The compound of formula (III') can be obtained by reacting a ketonecompound of the formula (IV'): ##STR6## wherein R¹, R³, R⁴, R⁵, R⁶, R⁷,R¹⁰, R¹¹, R¹², m and k are as defined above, with, for example,dimethyloxosulfonium methylide or dimethylsulfonium methylide in casen=1, in the presence of a diluent.

The ketone compound of formula (IV') can be obtained fromcyclopentanone-2-carboxylate and a corresponding substituted phenylhalide according to a known process (for example, Organic Syntheses,Vol. 45, 7, 1965).

The produced stereoisomers may be separated in each step. Separation ofthe stereoisomers can be accomplished by suitable means such ascrystallization or chromatography, preferably optical active columnchromatography.

Examples of the solvent used in the preparation of the compounds offormula (I) include ethers such as diethyl ether, diisopropyl ether andtetrahydrofuran; aromatic hydrocarbons such as benzene and toluene,containing ether; ethylene glycol; pyridine; and dimethylformamide. Thereaction is preferably carried out with stirring. After termination ofthe reaction, the obtained reaction mixture is extracted with an organicsolvent such as ethyl acetate, diethyl ether, chloroform, and benzene.The organic layer is separated, washed with water and dried, and thenthe solvent is distilled off under reduced pressure from the driedorganic layer. The thus obtained residue may be immediately applied tofurther reaction, but it is preferably purified by suitable purificationsuch as recrystallization or column chromatography before subjected tofurther reaction.

The present invention provide an aromatase-inhibiting compositioncharacterized by containing a compound selected from the groupconsisting of azole derivatives represented by the formula (I),including their stereoisomers and salts thereof. The stereoisomer may beused either singly or as a mixture of two or more of them. They may alsobe used in the form of a pharmaceutically acceptable salt.

The aromatase-inhibiting activity was measured basically according tothe method described in D. F. Covey et al: BBRC (I), 81-86, 1988. Thearomatase-inhibiting activity of the azole derivatives used in thepresent invention was evaluated in terms of 50% aromataseactivity-inhibiting concentration (IC₅₀) of the derivatives. The IC₅₀was not more than 1×10⁻⁸ M for all of the derivatives.

The azole derivatives according to the present invention are useful asaromatase inhibitors and especially useful for the treatment ofestrogen-dependent diseases such as breast cancer, benign breastdisease, uterine cancer, pancreatic carcinoma and Cushing's syndrome.

The acute toxicity (LD₅₀) of the azole derivatives of formula (I) asdetermined by subcutaneous injection to mice is above 300 mg/kg.

The azole derivative according to the present invention can beadministered through various routes such as oral, subcutaneous,intramuscular, intravenous, transdermal and rectal. It is usually usedin the form of pharmaceutical compositions with a pharmaceuticalacceptable carrier or diluent. For preparation of the compositions, theazole derivative (active ingredient) is usually mixed with a carrier,diluted with a carrier, or encapsulated in a carrier having a containerform such as capsule. The carrier as a diluent, may be a solid,semi-solid or liquid material which serves as a vehicle, excipient ormedium. Thus, the aromatase-inhibiting compositions of the presentinvention may be prepared into the form of tablet, pill, powder, elixir,emulsion, solution, syrup, suspension, aerosol, ointment, soft and hardgelatin capsules, suppository, sterile injectable solution and the like.

Examples of suitable carriers and diluents include lactose, dextrose,sucrose, sorbitol, mannitol, starch, calcium phosphate, calciumsilicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose,polyoxyethylene sorbitan mono-oleate, gelatin, syrup, methyl cellulose,methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearateand water. The composition may additionally contain lubricant, wettingagent, emulsifying agent, suspending agent, antiseptic, sweeteningagent, flavoring agent and the like.

For oral administration, for example, the azole derivative according tothe present invention may be admixed with carriers or diluents andprepared into powders, or molded into tablets or encapsulated in gelatincapsules. The mixtures can alternatively be dissolved in liquids such asan aqueous glucose solution, isotonic sodium chloride solution, sterilewater or the like.

The composition may be formulated in a unit dosage form which contains0.01 to 500 mg, preferably 0.1 to 300 mg of the azole derivative. Thecomposition is preferably formulated in a unit dosage form foradministration.

The azole derivative according to the present invention is effectiveover a wide dosage range. For example, dosages per day will normallyfall within the range of 0.005 to 100 mg/kg body weight. In thetreatment of human adults, a range of about 0.1 to about 40 mg/kg, insingle or divided doses, is preferred. However, it should be understoodthat the amount of the azole derivative actually administered will bedetermined by a skilled physician, in view of the relevant circumstancesincluding the age of the patient, the severity of the patient and theroute of administration. Therefore, the above dosage ranges are notintended to limit the scope of the invention in any way.

EXAMPLES

The following examples further illustrate the present invention.

Example 1

Preparation according to process A:

(1) Step (a): Preparation of ethyl 3,3-ethylenedioxy-4-methylpentanoate(VIII-1)

Ethyl 4-methyl-3-oxopentanoate (IX-1) (5.43 g), ethylene glycol (7.45 g)and p-toluenesulfonic acid (137.2 mg) were fed into an eggplant typeflask (500 ml). Benzene was poured into the mixed solution, and themixture was heated to reflux for 6 hours and the water was collectedthrough a Dean-Stark trap. After completion of the reaction wasconfirmed by thin-layer chromatography (TLC), the eggplant type flaskwas transferred onto an ice bath. A few drops of triethylamine wereadded to the reaction solution to neutralize it. Distilled water (50 ml)was added to the reaction solution and the mixture was extracted oncewith 100 ml of ethyl acetate and twice with 150 ml of ethyl acetate.

The extract was washed with saturated brine, dried over anhydrous sodiumsulfate and filtered, and the filtrate was concentrated under reducedpressure to obtain a crude product (6.84 g). The crude product waspurified by silica gel chromatography (Kieselgel 60, 100 g, φ=5.5 cm,n-hexane/ethyl acetate=6:1) to give the title compound (6.60 g).

Yield: 95.1%

¹ H-NMR (CDCl₃, δ ppm): 0.96 (s, 3H), 0.97 (s, 3H), 1.27 (m, 3H), 2.12(m, 1H), 2.67 (s, 2H), 3.96 (m, 2H), 4.02 (m, 2H) , 4.14 (m, 2H) .

IR (neat, cm⁻¹): 2990s, 2900m, 1740s, 1370w, 1185m.

(2) Step (b): Preparation of 3,3-ethylenedioxy-4-methyl-1-pentan-1-ol(VII-1)

Lithium aluminum hydride (950 mg) and dry tetrahydrofuran (THF, 20 ml)were stirred at room temperature for 10 minutes in an eggplant typeflask (100 ml). To the solution, the compound (VIII-1) (3.36 g) wasadded dropwise while cooled on an ice bath and the mixed solution wasstirred overnight at room temperature in an argon stream. Aftercompletion of the reaction was confirmed by TLC, distilled water (30 ml)was added to the reaction solution on an ice bath to terminate thereaction. The reaction solution was filtered through Celite 535 andextracted with ethyl acetate. The extract was dried over anhydroussodium sulfate, filtered and concentrated under reduced pressure. Theresulting crude product (2.27 g) was purified by silica gelchromatography (Kieselgel 60, 30 g, φ=3.0 cm, n-hexane/ethylacetate=2:1) to give the title compound (2.09 g) as a transparent oilysubstance.

Yield: 78.6%.

¹ H-NMR (CDCl₃, δ ppm): 0.94 (s, 3H), 0.95 (s, H), 1.95 (m, 3H),3.75 (t,2H), 4.00 (m, 4H).

IR (neat, cm⁻¹): 3450s, 2990s, 2920s, 1480m, 1390m, 1215m.

(3) Step (c): Preparation of 3,3-ethylenedioxy-4-methylpentylmethanesulfonate (VI-1)

Methanesulfonyl chloride (320.2 mg) was placed in an eggplant type flask(100 ml) on an ice bath, followed by dropwise addition of the compound(VII-1) (405 mg) and further addition of pyridine (2 ml). The mixedsolution was stirred in an argon stream. After completion of thereaction was confirmed by TLC, the reaction solution was allowed tostand for about three hours. HCl (1 N) was added dropwise to thereaction solution to adjust its pH to around 6. The reaction solutionwas extracted with ethyl acetate (10 ml×2) and the extract was washedwith saturated brine, dried over anhydrous sodium sulfate, filtered andconcentrated under reduced pressure to give a crude product (590.2 mg).

Yield: 97.3%.

¹ H-NMR (CDCl₃, δ ppm): 0.94 (s, 3H), 0.95 (s, 3H), 1.88 (m, 1H), 2.13(t, 2H), 3.01 (s, 3H), 3.95 (s, 4H), 4.33 (t, 2H).

IR (neat, cm⁻¹): 3650w, 3000s, 2920s, 1480m, 1360s, 1180s.

(4) Step (d) : Preparation of3,3-ethylenedioxy-1-(1H-imidazol-1-yl)-4-methylpentane (IV-1)

Sodium salt of imidazole (V-1) (323 mg) and methylformamide (DMF, 3 ml)were placed in an eggplant type flask (200 ml), and then the compound(VI-1) (569.1 mg) were added, and the mixture was stirred at 50° C. for3.5 hours on an oil bath. The oil bath was heated to 80° C. and themixture was stirred. After completion of the reaction was confirmed byTLC, the reaction solution was extracted with diethyl ether (50 ml×2).The extract was washed with saturated brine, dried over anhydrous sodiumsulfate, filtered and concentrated under reduced pressure. The resultingcrude product (494.9 mg) was purified by silica gel chromatography(Kieselgel 60, 10 g, φ=2.0 cm, n-hexane/ethyl acetate=1:1) to give thetitle compound (IV-1) (448.9 mg) as white crystals.

Yield: 86.4%.

¹ H-NMR (CDCl₃, δ ppm): 0.94 (s, 3H), 0.95 (s, 3H), 1.89 (m, 1H), 2.13(t, 2H), 3.99 (m, 6H), 6.92 (s, 1H), 7.05 (s, 1H), 7.48 (s, 1H).

IR (KBr, cm⁻¹): 3120s, 2990s, 1520s, 1460s, 1385s, 1235s, 1189s.

(5) Step (e): Preparation of 1-(1H-imidazol-1-yl)-4-methylpentan-3-one(III-1)

Sulfuric acid (1 ml) , THF (2 ml) and distilled water (1 ml) were placedin an eggplant type flask (200 ml), and the compound (IV-1) (211 mg) wasadded. The mixture was stirred overnight at room temperature. Aftercompletion of the reaction was confirmed by TLC, the reaction solutionwas neutralized with saturated sodium hydrogen carbonate (28 ml) on anice bath. The neutralized solution was extracted with ethyl acetate (50ml×2), and the extract was washed with saturated brine, dried overanhydrous sodium sulfate, filtered and concentrated under reducedpressure to give the title compound (III-1) (159.9 mg) as a yellowishoily substance.

Yield: 95.8%.

¹ H-NMR (CDCl₃, δ ppm): 1.05 (s, 3H), 1.06 (s, 3H), 2.53 (m, 1H), 2.90(t, 2H), 4.25 (t, 2H), 6.89 (s, 1H), 7.02 (s, 1H) , 7.47 (s, 1H).

IR (neat, cm⁻¹): 3420s, 3000s, 1715s, 1520m, 1475m, 1390m, 1235m, 1080m.

(6) Step (f): Preparation of(3R)-3-(4-chlorophenyl)-1-(1H-imidazol-1-yl)-4-methylpentan-3-ol (I-1)##STR7##

1-Bromo-4-chlorobenzene (II-1) (202 mg) and THF (1 ml) were placed in aneggplant type flask (200 ml), and n-butyl lithium (0.55 ml) was addeddropwise at -78° C. in an argon stream. One hour later, a dry THF (1 ml)solution of the compound (III-1) (146.0 mg) was added dropwise. Aftercompletion of the reaction was confirmed by TLC, the reaction mixturewas allowed to stand for 2 hours. The reaction was terminated withsaturated ammonium chloride (20 ml) of -53° C. and the reaction mixturewas heated to room temperature. Distilled water (3 ml) was added to thereaction solution and the solution was extracted with ethyl acetate (20ml×3). The extract was washed with saturated brine, dried over anhydroussodium sulfate, filtered and concentrated under reduced pressure. Thethus obtained crude product (204.3 mg) was purified by silica gelchromatography (Kieselgel 60, 10 g, φ=2.0 cm, n-hexane/ethylacetate/methanol=1:2:0.6) to give the title compound (I-1) (165.6 mg) asa white crystal.

Yield: 78.3%

¹ H-NMR (CDCl₃, δ ppm): 0.69 (d, 3H), 0.98 (s, 3H), 2.04 (m, 1H), 2.24(t, 1H), 2.33 (t, 1H) , 3.50 (m, 1H) , 3.95 (m, 1H), 6.79 (s, 1H), 7.00(s, 1H), 7.32 (d, 2H), 7.33 (s, 1H), 7.36 (d, 2H).

IR (KBr, cm⁻¹): 3200s, 2950s, 1520s, 1500s, 1230m, 1210m, 1090s.

(7) Step (f): Preparation of(3S)-3-(4-chlorobenzyl)-1-(1H-imidazol-1-yl)-4-methylpentan-3-ol (I-2)##STR8##

Magnesium (49 mg) and dry ether (2 ml) were placed in an eggplant typeflask (200 ml), and 4-chlorobenzyl chloride (II-1) (306 mg) was addeddropwise at room temperature in an argon stream. The mixed solution wasstirred for 30 minutes and refluxed on an oil bath of 40° C. 15 minuteslater, the solution was transferred onto an ice bath, and a dryTHF/ether (1:1) solution (1 ml) of the compound (III-1) (170 mg) wasadded dropwise. 30 minutes later, the reaction solution was returned toroom temperature and stirred overnight. After completion of the reactionwas confirmed by TLC, the reaction was terminated with saturatedammonium chloride (5 ml). Cold water (5 ml) was added to the reactionsolution and the solution was extracted with ethyl acetate (20 ml×2).The extract was washed with saturated brine, dried over anhydrous sodiumsulfate and concentrated under reduced pressure. The resulting crudeproduct (332.4 mg) was purified by silica gel chromatography (Kieselgel60, 15 g, φ=3.5 cm, n-hexane/ethyl acetate/methanol=1:1:0.3) to give thetitle compound (I-2) (208.6 mg) as white crystals.

Yield: 71.2%.

¹ H-NMR (CDCl₃, δ ppm): 0.98 (d, 3H), 1.04 (d, 3H), 1.78 (m, 2H), 2.00(m, 1H), 2.66 (d, 1H), 2.86 (d, 1H), 3.98 (m, 1H), 4.05 (m, 1H), 6.85(s, 1H), 7.03 (s, 1H), 7.15 (d, 2H), 7.30 (d, 2H), 7.44 (s, 1H).

IR (KBr, cm⁻¹): 3260s, 2980s, 2910s, 1520S, 1500s, 1470m, 1415m, 1235m,1095s.

Example 2

Preparation according to process B-1:

Preparation of1-(4-chlorobenzyl)-c-2-(1H-imidazol-1-ylmethyl)cyclopentan-r-1-ol (I-3)and its optical isomers (I-3 (+) and I-3 (-)) ##STR9##

Magnesium (1.48 g) and dry ether (15 ml) were placed and stirred in aneggplant type flask (200 ml) at room temperature in an argon stream. Tothe solution was added dropwise a dry ether (40 ml) solution of4-chlorobenzyl bromide (II-3) (12.5 g). The ether solution becamecloudy, and it was confirmed that the Grignard reagent began to form.While refluxing ether gently at room temperature, the remaining ethersolution of 4-chlorobenzyl bromide was added over a period of 25minutes. The solution was stirred for 2 hours and transferred onto awater bath of 5° to 10° C. To the solution, a dry THE (50 ml) solutionof 2-(1H-imidazol-1-ylmethyl)cyclopentan-1-on (III-3) (5.0 g) was addeddropwise over a period of 10 minutes, followed by 2-hour stirring of themixed solution. After disappearance of the starting materials wasconfirmed by TLC, the reaction solution was adjusted to pH 7 withaqueous HCl (1 N) solution, cooled and, after addition of a smallquantity of water, extracted with ethyl acetate. The organic layer waswashed with saturated brine, dried over anhydrous sodium sulfate andconcentrated under reduced pressure. The resulting crude product (8.83g) was crystallized with n-hexane/ethyl acetate (1:1) to form the crudecrystals (5.19 g), and these crude crystals (2 g) were purified bysilica gel column chromatography (270 g, acetone/n-hexane=1:1) to givethe title compound (I-3) (1.01 g) as white crystals.

Rf: 0.46 (ethyl acetate/n-hexane/methanol=5:3:1.5).

Mass spectrum: 290 (M⁺).

¹ H-NMR (CDCl₃, δ ppm): 1.47 (m, 1H), 1.58 (m, 2H), 1.80 (m, 3H), 2.11(m, 1H), 2.62 (s, 2H), 3.92 (dd, 1H), 4.16 (dd, 1H), 6.93 (s, 1H), 7.04(s, 1H), 7.12 (d, 2H), 7.28 (d, 2H), 7.50 (s, 1H).

IR (KBr, cm⁻¹): 3158s, 2990s, 1600w, 1585w, 1525s, 1508s.

The produced compound (I-3) was subjected to optical active columnchromatography (CHIRACEL OD, 20×250 mm, 10 μm, isopropanol/n-hexane=3:7,10 ml/min) to separate peak 1 and peak 2, from which the white crystals(170 mg) corresponding to the respective peaks were obtained,respectively. These crystals were recrystallized from methanol/water togive the crystals of 1-3 (-) (peak 1) (82.3 mg) and I-3 (+) (peak 2)(76.8 mg). I-3 (-) (peak 1)

Melting point: 172°-173° C. α!λ²⁵ : +31.3° (c=0.55, methanol). I-3 (+)(peak 2)

Melting point: 171°-172° C. α!λ²⁵ : +32.2° (c=0.57, methanol).

Further,1-(4-chlorobenzyl)-t-2-(1H-imidazol-1-ylmethyl)cyclopentan-r-1-ol (I-4)was separated by the column chromatography as white crystals.

Rf: 0.41 (ethyl acetate/n-hexane/methanol=5:3:1.5).

Mass spectrum: 290 (M+).

¹ H-NMR (CDCl₃, δ ppm): 1.42 (m, 2H), 1.73 (m, 3H), 1.93 (m, 1H), 2.37(m, 1H), 2.67 (d, 1H), 2.82 (d, 1H), 3.73 (dd, 1H), 4.30 (dd, 1H), 6.93(s, 1H), 7.05 (s, 1H), 6.99 (d, 2H), 7.30 (d, 2H), 7.46 (d, 1H).

IR (KBr, cm⁻¹): 3160s, 3000s, 1600w, 1522m, 1502s.

The following compounds were prepared in the similar way:

1-(Benzyl)-t-2-(1H-imidazol-1-ylmethyl)cyclopentan-r-1-ol (I-5), m.p.94°-98° C.

1-(4-Chlorobenzyl)-c-2-(1H-1,2,4-triazol-1-ylmethyl)cyclopentan-r-1-ol(I-6), m.p. 124°-126° C.

1-(2,4-Dichlorobenzyl)-c-2-(1H-1,2,4-triazol-1-ylmethyl)cyclopentan-r-1-ol(I-7), m.p. 104°-105° C.

1-(2,4-Difluorobenzyl)-c-2-(1H-1,2,4-triazol-1-ylmethyl)cyclopentan-r-1-ol(I-8), m.p. 99°-102° C.

1-(p-Biphenylmethyl)-c-2-(1H-1,2,4-triazol-1-ylmethyl)cyclopentan-r-1-ol(I-9), m.p. 150°-151° C.

1-(4-Chlorophenyl)-c-2-(1H-imidazol-1-ylmethyl)cyclopentan-r-1-ol(I-10), m.p. 128°-131° C.

1-(Phenyl)-c-2-(1H-imidazol-1-ylmethyl)cyclopentan-r-1-ol (I-11), m.p.92°-95° C.

1-(4-Chlorophenyl)-c-2-(1H-1,2,4-triazol-1-ylmethyl)cyclopentan-r-1-ol(I-12), m.p. 103°-105° C.

Example 3

Preparation according to process B-1:

Preparation of nitrate of1-(4-chlorobenzyl)-c-2-(1H-imidazol-1-ylmethyl)-cyclopentan-r-1-ol(I-13)

Compound (I-3) (0.1 g) was dissolved in THF and the solution was madeacidic (pH 2) with 70% nitric acid. Diisopropyl ether was added dropwiseto the solution to cause precipitation of white crystals. The crystalswere filtered out to give the title compound (I-13).

Mass spectrum: 354 (M⁺ +1), 290 (M⁺ -HNO3).

Example 4

Preparation according to process B-2:

Preparation of (3aR,9aR)-6-chloro-2,3,9,9a-tetrahydro-3,3-dimethyl-3a(1H)-(1H-1,2,4-triazol-1-ylmethyl)cyclopenta b! 1!benzopyran (I-14)##STR10##

To anhydrous DMF (30 ml) in an eggplant type flask (200 ml), sodiumhydride (670 ml, washed with anhydrous benzene) was added with stirringunder a helium atmosphere. To the solution was added 1H-1,2,4-triazole(V) (1.8 g) and the mixed solution was stirred at room temperature tillfoaming ceased. To the resulting solution, an anhydrous DMF (10 ml)solution of 7-(2-fluoro-4-chlorobenzyl-4,4-dimethyl-1-oxaspiro2,4!heptane (III'-14) (5.00 g) was added dropwise and the mixture wasstirred at 70° C. for 10 hours. The reaction solution was cooled, pouredonto ice-cold water and extracted with ethyl acetate. The organic layerwas washed with water and dried over anhydrous sodium sulfate and thesolvent was distilled away under reduced pressure. The residue waspurified by optical active column chromatography (CHIRACEL OD mfd. byDiacel Chem. Ind. Co., Ltd., isopropanol/n-hexane=3:7) to give the titlecompound (I-14) (1.5 g).

¹ H-NMR (CDCl₃, δ ppm): 0.80 (s, 3H), 1.12 (s, 3H), 1.12-3.07 (m, 7H),4.17 (d, 1H), 4.43 (d, 1H), 6.73 (b, 3H), 8.70 (s, 1H), 8.83 (s, 1H).

IR (KBr, cm⁻¹): 3060, 2940, 1595, 1570, 1480, 1130, 1020, 840, 740.α!λ²⁵ : -24.8° (c=0.71, methanol).

The following compounds were synthesized in the similar way:

(3aR,9aR)-6-Chloro-2,3,9,9a-tetrahydro-3a(1H)-(1H-imidazol-1-ylmethyl)-3,3-dimethylcyctopentab! 1!benzopyran (I-15), m.p. 108°-109° C.

(3aR,9aR)-6-Chloro-2,3,9,9a-tetrahydro-3a(1H)-(1H-imidazol-1-ylmethyl)cyclopentab! 1!benzopyran (I-16), m.p. 87°-88° C.

(3aR,9aR)-6-Chloro-2,3,9,9a-tetrahydro-3a(1H)-(1H-1,2,4-triazol-1-ylmethyl)cyclopentab! 1!benzopyran (I-17), m.p. 92°-94° C.

(3aR,9aR)-8-Fluoro-2,3,9,9a-tetrahydro-3a(1H)-(1H-1,2,4-triazol-1-ylmethyl)cyclopentab! 1!benzopyran (I-18), m.p. 149°-151° C.

The following compounds were also obtained in the similar way:

c-2-(4-Chlorobenzyl)-1-(1H-imidazol-1-ylmethyl)-4,4-dimethylcyclopentan-r-1-ol (I-19), m.p. 139°-140° C.

c-2-(4-Chlorobenzyl)-4,4-dimethyl-1-(1H-1,2,4-triazol-1-ylmethyl)cyclopentan-r-1-ol(I-20) , m.p. 84°-84.5° C.

Example 5

Acute toxicity

Acute toxicity was examined by subcutaneous administration to theICR-JCL mice. Each of the compounds (I-1) to (I-20) was suspended inolive oil, and a determined amount of each suspension was syringedsubcutaneously to the test mice. After administration, toxic symptoms ofthe mice were observed and LD₅₀ was determined from the number of deathsthat occurred during the period of 7 days after administration. LD₅₀ wasnot less than 300 mg/kg for all of the compounds tested.

Example 6

Aromatase-inhibiting activity

The aromatase-inhibiting activity of each compound was measuredaccording to the method of Covey et al: BBRC, 157 (1), 81-86, 1988.Radioactivity of a by-product H¹⁴ COOH released from a ¹⁴ C-labeledsubstrate by the action of aromatase was measured to radiometricallydetermine aromatase-inhibiting activity of the compounds. Thearomatase-inhibiting activity of the test compound is shown by 50%aromatase activity-inhibiting concentration (IC₅₀) of the compound.

An aromatase reaction system of the composition shown below in 67 mMphosphate buffer solution (0.5 ml) was incubated at 37° C. for 30minutes with shaking. Chloroform (5 ml) was added to the reactionsolution to stop the reaction. The reaction solution was vigorouslystirred and then centrifuged to recover the by-product (H¹⁴ COOH) of thearomatase reaction in the aqueous layer. The aqueous layer (0.1 ml) wascollected and mixed with a liquid scintillation cocktail (4 ml) tomeasure its radioactivity.

(1) Aromatase: Microsomes of human placenta (0.1 mg/ml, proteinconcentration).

(2) Substrate: 19-¹⁴ C! 4-Androstene-3,17-dione (1×10⁻⁶ M; 2 kBq/ml).

(3) Coenzymes: Nicotinamide adenine dinucleotide phosphate, reduced form(2×10⁻³ M), glucose-6-phosphate (4×10⁻³ M) and glucose-6-phosphatedehydrogenase (4 U/ml).

(4) Test compound

Radioactivity of the reaction system exclusive of (1) was supposed to bebackground radioactivity as 100% control of aromatase inhibition.Radioactivity of the reaction system exclusive of (4) was supposed to bethat of negative control as 0% of aromatase inhibition. Based on theresults of the measurements, a graph of the concentration of the testcompounds vs. the aromatase inhibiting activity was produced, and theIC50 was determined from the graph. 4-Hyroxyandrostenedione was used asa reference compound.

The results are shown in Table 1. The Compound Nos. correspond to thosegiven in the Examples 1-5.

                  TABLE 1                                                         ______________________________________                                               Comp. No.                                                                             IC.sub.50  (M)                                                 ______________________________________                                               I-1     1.0 × 10.sup.-8                                                 I-2     1.0 × 10.sup.-8                                                 I-3     4.0 × 10.sup.-8                                                 I-4     2.0 × 10.sup.-8                                                 I-5     5.0 × 10.sup.-8                                                 I-6     4.0 × 10.sup.-8                                                 I-7     1.0 × 10.sup.-8                                                 I-8     1.2 × 10.sup.-8                                                 I-9     1.3 × 10.sup.-8                                                 I-10    3.0 × 10.sup.-8                                                 I-11    3.5 × 10.sup.-8                                                 I-12    1.0 × 10.sup.-8                                                 I-13    5.5 × 10.sup.-8                                                 I-14    1.2 × 10.sup.-8                                                 I-15    4.5 × 10.sup.-8                                                 I-16    3.0 × 10.sup.-8                                                 I-17    3.0 × 10.sup.-8                                                 I-18    1.3 × 10.sup.-8                                                 I-19    6.5 × 10.sup.-8                                                 I-20    4.0 × 10.sup.-8                                                 Reference                                                                             2.0 × 10.sup.-8                                          ______________________________________                                    

Example 7

Aromatase inhibiting activity of optical isomers

The aromatase inhibiting activities (IC₅₀) of the optical isomers (I-3(-) and I-3 (+)) were determined in the same way as in Example 6. IC₅₀of I-3 (-) was 4.0×10⁻⁹ M and that of I-3 (+) was 8.0×10⁻⁸ M.

Example 8

Formulation example

    ______________________________________                                        Compound (I-1)           100    mg                                            Polyoxyethylene sorbitan mono-oleate                                                                   50     mg                                            Starch                   250    mg                                            ______________________________________                                    

The composition was mixed well and encapsulated in a capsule.

What is claimed is:
 1. A method of treating any one of breast cancer,benign breast disease, uterine cancer, pancreatic carcinoma, andCushing's syndrome, which comprises administering to a patient sufferingtherefrom an aromatase-inhibiting effective amount of a compoundselected from the group consisting of compounds of the formula (I):##STR11## including their stereoisomers and salts thereof, wherein R¹ isCl or Br; m is 0, 1 or 2; k and n each are independently 0 or 1; R³, R⁶and R⁷ each are H; R⁸ and R⁹ form a --CH₂ --C(R¹¹) (R¹²)-- bond, whereinR¹¹ and R¹² each are independently H or C₁₋₂ alkyl; Y is N or CH; andR¹⁰ and R² form an --O-- bond.
 2. A pharmaceutical compositioncomprising an aromatase-inhibiting effective amount of a compoundselected from the group consisting of compounds of the formula (I):##STR12## including their stereoisomers and salts thereof, wherein R¹ isCl or Br; m is 0, 1 or 2; k and n each are independently 0 or 1; R³, R⁶and R⁷ each are H; R⁸ and R⁹ form a --CH₂ --C(R¹¹) (R¹²)-- bond, whereinR¹¹ and R¹² each are independently H or C₁₋₂ alkyl; Y is N or CH; andR¹⁰ and R² form an --O-- bond, and a pharmaceutically acceptable carrieror diluent.
 3. A pharmaceutical composition according to claim 2,wherein the compound is a single stereoisomer.